Nejvíce citovaný článek - PubMed ID 30443352
Rayleigh-scattering microscopy for tracking and sizing nanoparticles in focused aerosol beams
The idea of using ultrashort X-ray pulses to obtain images of single proteins frozen in time has fascinated and inspired many. It was one of the arguments for building X-ray free-electron lasers. According to theory, the extremely intense pulses provide sufficient signal to dispense with using crystals as an amplifier, and the ultrashort pulse duration permits capturing the diffraction data before the sample inevitably explodes. This was first demonstrated on biological samples a decade ago on the giant mimivirus. Since then, a large collaboration has been pushing the limit of the smallest sample that can be imaged. The ability to capture snapshots on the timescale of atomic vibrations, while keeping the sample at room temperature, may allow probing the entire conformational phase space of macromolecules. Here we show the first observation of an X-ray diffraction pattern from a single protein, that of Escherichia coli GroEL which at 14 nm in diameter is the smallest biological sample ever imaged by X-rays, and demonstrate that the concept of diffraction before destruction extends to single proteins. From the pattern, it is possible to determine the approximate orientation of the protein. Our experiment demonstrates the feasibility of ultrafast imaging of single proteins, opening the way to single-molecule time-resolved studies on the femtosecond timescale.
- Publikační typ
- časopisecké články MeSH
The science of X-ray free-electron lasers (XFELs) critically depends on the performance of the X-ray laser and on the quality of the samples placed into the X-ray beam. The stability of biological samples is limited and key biomolecular transformations occur on short timescales. Experiments in biology require a support laboratory in the immediate vicinity of the beamlines. The XBI BioLab of the European XFEL (XBI denotes XFEL Biology Infrastructure) is an integrated user facility connected to the beamlines for supporting a wide range of biological experiments. The laboratory was financed and built by a collaboration between the European XFEL and the XBI User Consortium, whose members come from Finland, Germany, the Slovak Republic, Sweden and the USA, with observers from Denmark and the Russian Federation. Arranged around a central wet laboratory, the XBI BioLab provides facilities for sample preparation and scoring, laboratories for growing prokaryotic and eukaryotic cells, a Bio Safety Level 2 laboratory, sample purification and characterization facilities, a crystallization laboratory, an anaerobic laboratory, an aerosol laboratory, a vacuum laboratory for injector tests, and laboratories for optical microscopy, atomic force microscopy and electron microscopy. Here, an overview of the XBI facility is given and some of the results of the first user experiments are highlighted.
- Klíčová slova
- European XFEL (EuXFEL), XBI Laboratory, coherent diffractive imaging (CDI), free-electron lasers (XFELs), sample preparation and characterization, serial femtosecond crystallography (SFX), single-particle imaging (SPI), structural biology, time-resolved experiments,
- Publikační typ
- časopisecké články MeSH
Aerosol nanoparticle injectors are fundamentally important for experiments where container-free sample handling is needed to study isolated nanoparticles. The injector consists of a nebuliser, a differential pumping unit, and an aerodynamic lens to create and deliver a focused particle beam to the interaction point inside a vacuum chamber. The tightest focus of the particle beam is close to the injector tip. The density of the focusing carrier gas is high at this point. We show here how this gas interacts with a near infrared laser pulse (800 nm wavelength, 120 fs pulse duration) at intensities approaching 1016 Wcm-2. We observe acceleration of gas ions to kinetic energies of 100s eV and study their energies as a function of the carrier gas density. Our results indicate that field ionisation by the intense near-infrared laser pulse opens up a plasma channel behind the laser pulse. The observations can be understood in terms of a Coulomb explosion of the created underdense plasma channel. The results can be used to estimate gas background in experiments with the injector and they open up opportunities for a new class of studies on electron and ion dynamics in nanoparticles surrounded by a low-density gas.
- Publikační typ
- časopisecké články MeSH
The possibility of imaging single proteins constitutes an exciting challenge for x-ray lasers. Despite encouraging results on large particles, imaging small particles has proven to be difficult for two reasons: not quite high enough pulse intensity from currently available x-ray lasers and, as we demonstrate here, contamination of the aerosolized molecules by nonvolatile contaminants in the solution. The amount of contamination on the sample depends on the initial droplet size during aerosolization. Here, we show that, with our electrospray injector, we can decrease the size of aerosol droplets and demonstrate virtually contaminant-free sample delivery of organelles, small virions, and proteins. The results presented here, together with the increased performance of next-generation x-ray lasers, constitute an important stepping stone toward the ultimate goal of protein structure determination from imaging at room temperature and high temporal resolution.
